COVID-19 in immunocompromised patients after hematopoietic stem cell transplantation: a pilot study

Data on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in patients at early stage of immune reconstitution after hematopoietic stem cell transplantation (HSCT) are limited. In the present study, we retrospectively investigated the incidence and clinical features of SARS-CoV-2 infection in patients who underwent HSCT in 2022. Patients (allo-HSCT, n = 80; auto-HSCT, n = 37) were consecutively included in the study. The SARS-CoV-2 infection rate was 59.8%, and the median interval of HSCT to coronavirus disease 2019 (COVID-19) was 4.8 (range: 0.5–12) months. Most patients were categorized as mild (41.4%) or moderate (38.6%), and 20% as severe/critical. No deaths were attributable to COVID-19. Further analysis showed that lower circulating CD8+ T-cell counts and calcineurin inhibitor administration increased the risk of SARS-CoV-2 infection. Exposure to rituximab significantly increased the probability of severe or critical COVID-19 compared with that of mild/moderate illness (P < .001). In the multivariate analysis, rituximab use was associated with severe COVID-19. Additionally, COVID-19 had no significant effect on immune reconstitution. Furthermore, it was found that Epstein–Barr virus infection and rituximab administration possibly increase the risk of developing severe illness. Our study provides preliminary insights into the effect of SARS-CoV-2 on immune reconstitution and the outcomes of allo-HSCT recipients.


INTRODUCTION
The global coronavirus disease 2019 (COVID-19) pandemic that originated due to infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to pose daunting challenges, especially to vulnerable individuals, including cancer survivors.SARS-CoV-2 causes acute respiratory infections followed by hyperinflammation in a subset of patients who develop severe disease.As of 2023, the global death toll exceeded six million. 1A comprehensive metaanalysis indicated notably higher mortality rates in patients with hematological malignancies than in those with solid tumors (odds ratio = 1.64). 2,3Similarly, immune deficiency is expected to have a prolonged effect on the progression and outcome of SARS-CoV-2 infection. 4In a prospective cohort of patients with aplastic anemia, the percentage of patients with severe/critical COVID-19 was similar to that of the general population (0.6% vs 0.5%, respectively, P > .05) 5 but was substantially lower than that of patients with hematological malignancies. 6However, uncertainty surrounds the outcomes of SARS-CoV-2 infection in transplant recipients with underlying hematologic malignancies.These individuals are characterized by profound immune dysregulation caused by conditioning, acute/chronic graft-vs-host disease (GvHD), and long-term immunosuppressive therapy, 7 rendering them particularly susceptible to SARS-CoV-2 infection.Therefore, the Worldwide Network for Blood and Marrow Transplantation and the Center for International Blood and Marrow Transplant Research have issued guidelines to protect hematopoietic stem cell transplantation (HSCT) recipients and donors during the COVID-19 pandemic. 8However, their susceptibility to infection persists 9,10 and adequate data on the outcomes of transplant recipients with COVID-19 are scarce.
In December 2022, the worldwide spread of SARS-CoV-2 became notable, exposing all HSCT recipients to infection risk.The first year after transplantation is characterized by cellular and humoral immunodeficiencies and greater susceptibility to infection.In the present study, we aimed to characterize the clinical characteristics and outcomes of COVID-19 in HSCT recipients within the first year after HSCT during the first surge of the pandemic.By evaluating infections and outcomes after SARS-CoV-2 exposure, our study provides preliminary evidence for optimizing the prevention and control of potential SARS-CoV-2 infections among transplant recipients in the future.

Patients
This retrospective study was based on the transplant databases of Shanghai Ruijin Hospital and Shanghai Liquan Hospital.Consecutive hematologic patients receiving HSCT from January 1, 2022, to December 31, 2022, were screened.The eligibility criteria were as follows: age ≥ 16 years and available results of molecular diagnostic tests (real-time polymerase chain reaction) and/or antigen detection of SARS-CoV-2.The exclusion criteria were as follows: incomplete medical information and lack of follow-up data.The final follow-up was conducted on January 31, 2023.The study was approved by the institutional review board of each participating hospital (approval number: 2021-173) and conducted in accordance with the Declaration of Helsinki.

Transplant regimens, GvHD prophylaxis, and infection prophylaxis
The protocols for the preconditioning regimen, GvHD prophylaxis and treatment, and infection prophylaxis have been reported in detail. 11,123.Definition SARS-CoV-2 infection was diagnosed using molecular diagnostic tests and/or antigen detection.COVID-19 severity was classified according to the international recommendations of the National Institutes of Health: asymptomatic or presymptomatic infection (positive for SARS-CoV-2 virology test without clinical symptoms), mild (upper respiratory symptoms), moderate (acute lower respiratory tract infection, but no hypoxemia), severe (hypoxia ≤ 93% without supplemental oxygen), or critical (admission to intensive care unit).Patients were followed up for a minimum of 30 days after SARS-CoV-2 infection.Cytomegalovirus (CMV) viremia and Epstein-Barr virus (EBV) viremia were detected in the plasma samples at any level of CMV/EBV DNA.CMV disease was diagnosed according to the established criteria.13 Mixed infection was defined as the co-occurrence of SARS-CoV-2 and other pathogens in the same infection episode and/or different infection sites involved in the same episode.

Data acquirement
The investigators at each hospital used the institutional electronic medical records of clinical databases to obtain the required information.Collected data included patient demographics, diagnoses, transplant regimens, and clinical outcomes (relapse, mortality, and survival).The clinical data were collected using an electronic questionnaire.All data were independently reviewed by 2 experienced transplantation physicians to ensure the accuracy of the results.

Statistical analysis
Frequencies and percentages were used to describe patient characteristics.Univariate and multivariate analyses of the clinical, laboratory, and therapeutic variables associated with outcomes were performed using logistic regression models.For multivariate analysis, variables with parameter estimates with P ≤ .10 in the univariate analysis were included.Univariate and multivariate Cox regression analyses were performed to determine the effect of potential prognostic factors on the clinical outcomes.Two-sided exact P values were reported and P ≤ .05 was considered statistically significant.Statistical analyses were performed using Statistical Package for the Social Sciences version 26 (SPSS Inc.; IBM, Armonk, New York).

Mixed infection in patients with SARS-CoV-2 infection
Mixed infections were diagnosed in 32 patients (45.7%), with bacteria being the most common pathogen (n = 32, 45.7%) followed by fungi (n = 19, 27.1%).Patients in the mixed infection group were more likely to develop severe or critical disease than patients in the non-mixed infection group (43.9% vs 0%, respectively, P < .001).Compared with that in the severe COVID-19 patients without mixed infection, the duration of infection was markedly longer in the mixed infection group (13 days [range: 7-40 days] vs 9 days [5-37 days], respectively, P = .03).Moreover, CMV disease, EBV reactivation, and rituximab use were more commonly observed in the mixed infection group (Table 2).

Comparison of mild/moderate and severe/critical infections
Next, we compared the clinical characteristics of patients with mild/moderate and severe/critical SARS-CoV-2 infections.

Immune reconstitution
Immune reconstitution analysis was conducted in 58 of the 80 allo-HSCT patients.The results showed that lower numbers of CD8 + T cells were associated with increased SARS-CoV-2 infection (505/μL vs 1045/μL, respectively, P = .033,Table 3).The other lymphocyte subsets did not differ significantly between the infected and non-infected groups.The distribution of lymphocyte subsets before SARS-CoV-2 exposure did not relate to COVID-19 severity.
Recovery of the immune system after HSCT is highly dynamic.To evaluate the long-term effect of SARS-CoV-2 infection on immune reconstitution, we tracked the lymphocyte subsets of these patients 3 and 6 months after SARS-CoV-2 exposure.There were no differences in the absolute counts of CD3 + , CD4 + , CD8+, NK, and B cells between the infected and non-infected groups at any time point.These results indicate that SARS-CoV-2 has a subtle effect on immune reconstitution (Fig. 5).

DISCUSSION
To date, the global number of detected COVID-19 cases is more than 35 million. 14Most studies conducted in the prevaccine era reported mortality rates higher than 40% among patients with hematological malignancies. 15,16However, recent studies have reported an average mortality rate of 4% (1%-20%).The uneven burden of COVID-19 across different populations is a hallmark of this pandemic, 17 and patients with hematologic malignancies show prolonged viral shedding and higher mortality than the general population and even patients with solid tumors. 18Moreover, delayed immunization in patients undergoing HSCT increases the risk of life-threatening sequelae and adverse outcomes of SARS-CoV-2 infection. 19New SARS-CoV-2 variants have emerged sequentially, presenting distinct virulence, transmissibility, and host immune responses. 20The recently dominant Omicron variants in China are marked by increased transmissibility but decreased lethality. 21SARS-CoV-2 Omicron variants have evolved to evade neutralizing antibodies elicited by previous infections and vaccination, 22 presenting a new challenge for HSCT patients.Therefore, it is imperative to perform a retrospective analysis to understand the recent implications of COVID-19 on patients undergoing HSCT.
In this study, we assessed the incidence and outcomes of COVID-19 in 117 consecutive HSCT recipients.There were no statistically significant differences in age, sex, primary disease, or HSCT characteristics between the infected and non-infected indicating that these factors did not increase the risk of SARS-CoV-2 infection in HSCT recipients.Immunosuppression may increase the risk of SARS-CoV-2 infection, particularly in severe illnesses. 23Immunosuppressive agents, including cyclosporine and tacrolimus, were maintained in 31.2% of the patients before SARS-CoV-2 exposure.Although little evidence of immunosuppression as a significant risk factor for SARS-CoV-2 infection and severe disease was observed in a larger cohort of immunosuppressed patients with liver transplants 24 and inflammatory bowel disease, 25 our data showed that patients who received CNIs were at an increased risk of SARS-CoV-2 infection (41.4% vs 21.3%, respectively, P = .028).The role of TKI as a prominent risk factor for COVID-19 has been established in a cohort of patients with chronic myeloid leukemia cohort. 26However, there is no evidence that TKI administration negatively affects outcomes for COVID-19 patients. 27In our study, TKIs were used as maintenance therapy for 6 patients after allo-HSCT, 5 of whom developed mild/moderate SARS-CoV-2 infection.However, the influence of TKIs on COVID-19 susceptibility needs to be verified in larger populations.
The clinical profile and overall course of COVID-19 in our study were similar to those in larger cohorts of patients with hematological malignancies who underwent HSCT. 28,29Overt symptoms upon initial assessment commonly included fever, cough, and fatigue, a trend evident across all patient groups.Nevertheless, our study found that severe/critical illness accounted for 12% of all infected episodes, which was higher than the proportions reported in other studies on HSCT populations. 28This difference could be attributed to the higher proportion of patients who underwent allo-HSCT (68.4%) in our study, which is consistent with previous results of allo-HSCT patients. 30maging is not routinely indicated for COVID-19 in asymptomatic individuals or patients with mild symptoms unless they are at risk of disease progression.In this study, 54 patients underwent lung CT.Typically, radiologic findings are contingent upon the disease stage, 31 with frequent radiographic findings in COVID-19 pneumonia, including multifocal consolidation or ground-glass opacity, which may progress to pneumonia. 32f the patients, 33% showed consolidation and/or groundglass opacities, with no progression to organizing pneumonia.These results confirmed that patients infected with the Omicron variant (B.1.1.529)had a lower risk of pneumonia and clinical severity than those infected with alternative variants, in agreement with previous reports. 33Viral load in the lower respiratory tract may persist at a higher level for a prolonged period.Thus, BALF is a more reliable sample for immunocompromised patients compared with nasopharyngeal swab. 34ore than half the patients in our cohort (n = 49) were monitored at an outpatient clinic.Although several reported risk factors associated with disease severity were noted, they did not show a substantial effect in the present study, likely owing to the small size of our group.6][37][38][39] There are several possible explanations for this discrepancy.First, there have been considerable advances in COVID-19 vaccinations and treatments, including the advent of antiviral medications, which have greatly reduced the incidence of severe or critical infection. 20,40,41In addition, Omicron variants exhibit reduced pathogenicity, resulting in weaker inflammatory responses than the other variants. 42Cohen et al 43 reported that the probability of severe illness reduced by 73% during an Omicron-dominated wave.
Most patients undergoing HSCT require scheduled outpatient visits, which could potentially expose them to a variety of pathogens, including SARS-CoV-2.Further exploration of the vulnerability of immunodeficient patients to viral infections remains a priority.The immunological pathogenicity of SARS-CoV-2 is complicated, given its virulence and lack of temporal coordination between innate and adaptive immune responses. 44B cell-depleting agents, such as rituximab, which target the CD20 marker on B cells, are widely used for B cell hematological malignancies, EBV reactivation after allo-HSCT, and post-transplant lymphoproliferative disorders. 45However, the safety of rituximab in the context of COVID-19 is still under review. 46B-cell depletion compromises antiviral immunity, increases the risk of reactivation, and reduces viral clearance.However, it may be beneficial in specific scenarios as a mechanism for regulating adaptive host immune responses in patients with COVID-19.Severe infection and prolonged hospital stay increase the risk of morbidity, mortality, and infection-related sequelae, as observed in a study of patients with inflammatory rheumatic and musculoskeletal diseases treated with rituximab. 47In the present study, we identified    Chin Assoc of Blood Sci rituximab as potential risk factor for severe COVID-19, which has not been reported previously.Additionally, the median interval between COVID-19 exposure and the last rituximab infusion was shorter in severe/critical patients than in those with mild/ moderate infection.However, the exact causal relationship needs to be verified in a larger cohort.
Our study is limited by its retrospective design and relatively short follow-up period.Furthermore, cytokines have an important effect on the reconstitution of lymphocyte subsets and immune responses after SARS-CoV-2 infection.Due to the retrospective nature of our study, we could not collect enough data to thoroughly analyze the effect of cytokines on illness.Nonetheless, we believe that our study offers a comprehensive examination specific to HSCT recipients with COVID-19.
In conclusion, our study highlights the outcomes of SARS-CoV-2 infection among HSCT patients during the Omicron variant wave and provides new insights into the risk factors associated with COVID-19 severity.Our data suggest that EBV reactivation and rituximab use would increase the risk of developing severe disease.Ongoing surveillance of immune reconstitution is critical to evaluate the long-term impact of SARS-CoV-2 infection in HSCT recipients.

AUTHOR CONTRIBUTIONS
X.H., S.L., and S.Z.designed the study; Z.Z. and J.H. wrote the manuscript.Z.Z., J.H., L.W., and Z.P. were involved in collecting, analyzing, or interpreting research data and writing the manuscript.C.J. and J.H. analyzed research data.
This work was supported by the National Key Research and Development Program of China (no.2022YFC2502600), the

Table 2
The comparison of coinfection and non-coinfections of SARS-CoV-2.